Centrifugation

Objectives of Centrifugation

1. Separation of Components: To separate particles based on their size, shape, density, viscosity of the medium, and rotor speed.
2. Purification: To purify biological samples, such as proteins, nucleic acids, cells, and sub-cellular organelles.
3. Concentration: To concentrate particles, cells, or macromolecules from a solution.
4. Isolation: To isolate specific components from a complex mixture for further analysis or use.
5. Clarification: To remove suspended particles from a solution, thereby clarifying it.

Principle of Centrifugation

• Centrifugation operates based on the principle of sedimentation, where a force greater than gravity is applied to accelerate the settling rate of particles in a sample.
• When a sample is spun at high speed in a centrifuge, particles with different densities move at different rates:

Centrifugal Force:

Particles are subjected to a centrifugal force,

$F_c = m \cdot r \cdot \omega^2$

• m is the mass of the particle,
• r is the radius of rotation,
• ω is the angular velocity.

Sedimentation Rate:

• The rate at which particles sediment depends on their mass, the centrifugal force, and the frictional resistance they encounter from the medium they are suspended in.
• Particles with higher density or larger size sediment faster towards the bottom of the tube, forming a pellet, while lighter or smaller particles remain in the supernatant.

Applications of Centrifugation

1. Biological and Clinical Laboratories:

   1. Cell Separation: Isolating cells from blood or other body fluids.
   2. Fractionation of Subcellular Organelles: Separating organelles like nuclei, mitochondria, lysosomes, and ribosomes.
   3. DNA and RNA Purification: Isolating nucleic acids from cell lysates.
   4. Protein Purification: Separating proteins based on size and density.

2. Pharmaceutical Industry:

   1. Drug Development: Purifying drugs, vaccines, and other pharmaceutical products.
   2. Quality Control: Ensuring the purity and concentration of pharmaceutical preparations.

3. Food and Beverage Industry:

   1. Milk Processing: Separating cream from milk.
   2. Wine Clarification: Removing yeast and other particulates from wine.

4. Environmental Science:

   1. Water and Wastewater Treatment: Removing suspended solids and microorganisms from water samples.
   2. Soil Analysis: Separating soil components for analysis of contaminants.

5. Industrial Applications:

   1. Nanoparticle Separation: Isolating nanoparticles for use in various industrial processes.
   2. Oil and Gas: Purifying lubricants and other fluids.

Types of Centrifugation

1. Differential Centrifugation:

   • Based on differences in sedimentation rates of particles.
   • Commonly used for separating cells, organelles, and macromolecules.

2. Density Gradient Centrifugation:

   • Uses a gradient medium (e.g., sucrose or cesium chloride) to separate particles based on density.
   • Can be further classified into isopycnic (buoyant density) and rate-zonal (size and shape) centrifugation.

3. Ultracentrifugation:

   • Uses extremely high speeds (over 100,000 rpm) for separating very small particles like viruses, ribosomes, and macromolecules.
   • Requires special equipment and is often used in research settings.

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